Ubiquitin is a highly conserved 76-amino acid protein implicated in the function of quite different vital cellular processes. In the present study, we cloned and sequenced a polyubiquitin gene from Tuber borchii (Ubil) that is organised in four tandem repeats, with two C-terminal extension amino acids, serine and leucine. Two introns of 116 bp and 55 bp in length were detected in the first and second repeats, respectively. The Ubil gene is highly expressed in mycelium and is less expressed in the ripe fruiting body. Southern and Northern blot analyses revealed a second form of the ubiquitin gene.